Vaccines are typically used to protect humans and veterinary animals from infectious diseases caused by bacteria, viruses and parasitic organisms. The antigens used in vaccines may be any variety of agents but are typically composed of killed pathogenic organisms, pathogenic organisms which are alive but modified or attenuated, proteins, recombinant proteins or fragments thereof. Whatever the source of the antigen, it is often necessary to add an adjuvant to enhance the host immune response to the antigen.
Adjuvants are used to accomplish two objectives: they slow the release of the antigens from the injection site and they stimulate the immune system.
The first adjuvant reported in the literature was Freund's Complete Adjuvant (FCA). FCA contains a water-in-oil emulsion and extracts of mycobacterium. The mycobacterium extracts provide immunostimulatory molecules in a crude form. The water-in-oil emulsion acts to create a depot effect where the antigens are slowly released. Unfortunately FCA is poorly tolerated and it can cause uncontrolled inflammation. Since the discovery of FCA over 80 years ago efforts have been made to reduce the unwanted side effects of adjuvants.
Glycolipid analogues comprising a new class of compounds having adjuvant properties are now known. U.S. Pat. No. 4,855,283, (hereinafter '283) discloses the synthesis of glycolipid analogues, including N-glycosylamides, N-glycosylureas, N-glycosylcarbamates, and specifically: N-(2-deoxy-2-L-leucylamino-β-D-glucopyranosyl)-N-octadecyldodecanamide acetate (known as Bay R1005®, O Lockhoff, Angew. Chem. Int. Ed. Engl. (1991) 30:1611-1620. The compounds described in the '283 patent are particularly suitable for use as adjuvants.
Glycolipid adjuvant formulations need to be easy to manufacture and stable when stored for long periods of time without showing flocculation of the lipid component. The non-acetate forms of the glycolipid amides or glycosylamides are highly insoluble and typically flocculate out of solution upon storage either at room or lower temperatures.
The solutions and adjuvants comprising glycosylamides provided here show little flocculation and are quite stable. They are easy to manufacture and can be prepared on a commercial scale. The liquid glycolipid adjuvant formulations can be used as a diluent to rehydrate a lyophilized antigen preparation. Methods to test the stability of these formulations in real time and through accelerated stability testing protocols are also provided.